Production method for radiolabeled aryl compound

ABSTRACT

The invention relates to a method of producing the radiolabeled aryl compound (I) Ar—X, or a salt thereof, wherein X is 211At, 210At, 123I, 124I, 125I, or 131I. The method involves reacting the aryl boronic acid compound (II) Ar—Y, or a salt thereof, wherein Y is a borono group (—B(OH)2) or an ester group thereof, with a radionuclide selected from 211At, 210At, 123I, 124I, 125I and 131I, in the presence of an oxidizing agent selected from an alkali metal iodide, an alkali metal bromide, N-bromosuccinimide, N-chlorosuccinimide and hydrogen peroxide, in water.

CROSS-REFERENCE TO RELATED APPLICATIONS

This patent application is the U.S. national phase of InternationalPatent Application No. PCT/JP2018/030006, filed on Aug. 3, 2018, whichclaims the benefit of Japanese Patent Application No. 2017-151632, filedAug. 4, 2017, the disclosures of which are incorporated herein byreference in their entireties for all purposes.

TECHNICAL FIELD

The present invention relates to a production method of radiolabeledaryl compound applicable to RI internal therapy or diagnosis for cancer.

BACKGROUND ART

RI internal therapy or diagnosis for cancer by use of radionuclides thatemit α-ray, β-ray, γ-ray and the like utilizes specific bindings ofradionuclide-labeled drugs to target molecules, i.e., moleculesspecifically expressed or overexpressed in cancer cells, and it has beenapplied in a clinical practice. For example, Na¹³¹I has been applied totherapy for thyroid cancer, and ²²³RaCl₂ has been applied to therapy forprostate cancer bone metastasis.

Application of ²¹¹At, one of radionuclides, is expected as new RIinternal therapy for cancer (e.g., 4-²¹¹At-L-phenylalanine (Non-PatentDocument 1), Na²¹¹At, etc.). ²¹¹At is a radionuclide produced by anaccelerator such as cyclotron and the like, and has a short half-life of7.2 hours. Therefore, a sequence of processes containing production of²¹¹At, labelling of a drug with ²¹¹At, formulation of the drug,administration of the drug to patient with cancer, and RI internaltherapy by the drug should be promptly carried out. In particular, sincethe labelling and the subsequent formulation should be easily carriedout in a short time, the formulation is desirably carried outimmediately after the labelling. Moreover, since the labeled drug is tobe formulated into an injection for intravenous administration, thelabeling is desirably carried out without using toxic reagent, under anorganic solvent-free condition composed only of water, and the like, interms of prompt formulation after the labelling. ¹²³I for diagnosis alsohas a short half-life of 13.23 hours, and therefore, the labelling andthe subsequent formulation should be easily carried out in a short time,as in the case in ²¹¹At.

Non-Patent Document 1 discloses that 4-²¹¹At-L-phenylalanine can beapplied to RI internal therapy for brain tumor, and a precursor,N-Boc-4-tributylstannyl-L-phenylalanine is produced in a radiochemicalyield of 35-50%, by electrophilic destannylation according to a methoddescribed in Non-Patent Document 2. However, since the precursor is anN-Boc form, it cannot be dissolved in a solvent composed only of water,and use of an organic solvent for dissolution requires evaporation. Inaddition, the above method requires de-Boc step after the electrophilicdestannylation. Moreover, Non-Patent Document 2 does not disclosespecific methods for the de-Boc step, only specifically discloseslabelling of 4-iodo-L-phenylalanine with ²¹¹At by halogen exchangereaction in the presence of CuSO4, SnSO4 and an acid, at 120° C. for 60minutes. The method requires removal of toxic Cu and Sn, and thereaction at 120° C. for 60 minutes is not an easy method in a shorttime. The labelling by the above-mentioned methods is not desirable, andlabelling and the subsequent formulation cannot be easily carried out ina short time. Moreover, the radiochemical yield is low andmost-unsatisfactory.

Non-Patent Document 3 discloses that aryl boronic acid or an esterthereof is labeled with Na¹²³I by electrophilic substitution reaction inthe presence of 1,10-phenanthroline and a Cu catalyst such as Cu₂O,Cu(OCOCF₃)₂ and the like, at 80° C., in water/methanol. However, themethod is not desirable in terms of use of methanol and a Cu catalyst,and reaction at high temperature. In addition, the radiochemical yieldis at most 87% and unsatisfactory. Moreover, the document also disclosesthat aryl boronic acid or an ester thereof is labeled with Na¹²³I byelectrophilic substitution reaction in the presence of chliramine-T, inwater/tetrahydrofuran. However, the method is not desirable in terms ofuse of tetrahydrofuran, and the method cannot be applied to an electrondeficient arene.

Patent Documents 1 to 4 discloses the aryl trialkyltin is labeled withNa¹²³I, Na²¹¹At and the like by electrophilic destannylation reaction.However, the reactions are not desirable because of the use of anorganic solvent and toxic Sn.

DOCUMENT LIST Patent Document

-   Patent Document 1: U.S. Pat. No. 4,826,672-   Patent Document 2: U.S. Pat. No. 5,077,035-   Patent Document 3: JP 2001-503412-   Patent Document 4: JP 2009-521469

Non-Patent Document

-   Non-Patent Document 1: Nuklearmedizin, 2013, vol. 52, pp. 212-21-   Non-Patent Document 2: Applied Radiation and Isotopes, 2010, vol.    68, pp. 1060-1065-   Non-Patent Document 3: Chem. Commun., 2016, vol. 52, pp. 13277-13280

SUMMARY OF THE INVENTION Problems to be Solved by the Invention

The present invention aims to produce a radiolabeled aryl compound by amethod that enables an easy labelling with a high radiochemical yield ina short time, and that enables formulation immediately after thelabelling.

Means of Solving the Problems

The present inventors have conducted intensive studies in an attempt tosolve the above-mentioned problems and found that, by the followingmethod, a radiolabeled aryl compound can be easily produced in a highradiochemical yield in a short time, and the formulation can be carriedout immediately after the labelling, which resulted in the completion ofthe present invention.

Accordingly, the present invention provides the following.

-   [1] A method of producing a radiolabeled aryl compound represented    by the formula (I):    Ar—X   (I)    wherein-   Ar is a C₆₋₁₄ aryl group optionally having substituent(s), and X is    ²¹¹At, ²¹⁰At, ¹²³I, ¹²⁴I, ¹²⁵I or ¹³¹I,-   or a salt thereof (hereinafter, sometimes to be referred to as    radiolabeled aryl compound (I)), which comprises reacting an aryl    boronic acid compound represented by the formula (II):    Ar—Y   (II)    wherein-   Ar is as defined above, and-   Y is a borono group (—B(OH)₂) or an ester group thereof,-   or a salt thereof (hereinafter, sometimes to be referred to as    radiolabeled aryl compound (II)), with a radionuclide selected from    ²¹¹At, ²¹⁰At, ¹²³I, ¹²⁴I, ¹²⁵I or ¹³¹I, in the presence of an    oxidizing agent selected from an alkali metal iodide, an alkali    metal bromide, N-bromosuccinimide, N-chlorosuccinimide and hydrogen    peroxide, in water.-   [2] The method according to the above-mentioned [1], wherein the    reaction is carried out in an organic solvent-free system.-   [3] The method according to the above-mentioned [1] or [2], wherein    the reaction is carried out at room temperature.-   [4] The method according to any of the above-mentioned [1] to [3],    wherein the radionuclide is ²¹¹At or ²¹⁰At, and the oxidizing agent    is selected from sodium iodide, sodium bromide, N-bromosuccinimide,    N-chlorosuccinimide and hydrogen peroxide.-   [5] The method according to any of the above-mentioned [1] to [3],    wherein the radionuclide is ¹²³I, ¹²⁴I, ¹²⁵I or ¹³¹I, and the    oxidizing agent is selected from N-bromosuccinimide and    N-chlorosuccinimide.-   [6] The method according to any of the above-mentioned [1] to [5],    wherein Y is a borono group (—B(OH)₂).-   [7] The method according to any of the above-mentioned [1] to [6],    wherein the substituent of the C₆₋₁₄ aryl group optionally having    substituent(s) represented by Ar is a group capable of binding    specifically to a target molecule.-   [8] The method according to the above-mentioned [7], wherein the    target molecule is an antigen, a transporter, a receptor, an enzyme    or a gene, which is specifically expressed or overexpressed in a    cancer cell.-   [9] The method according to any of the above-mentioned [1] to [8],    wherein Ar is-   a group represented by the formula:

wherein

-   R² is a halogen atom,-   m is 0 or 1,-   n is 0 or an integer of 1 to 4, and-   * is a binding site to X or Y, or-   a group derived from a peptide having a partial structure    represented by the formula:

wherein

-   R² is a halogen atom,-   m is 0 or 1,-   n is 0 or an integer of 1 to 4, and-   * is a binding site to X or Y.-   [10] The method according to any of the above-mentioned [1] to [8],    wherein Ar is-   a group represented by the formula:

wherein

-   R³ is a hydrogen atom or a halogen atom,-   m is 0 or 1, and-   * is a binding site to X or Y, or-   a group derived from a peptide having a partial structure    represented by the formula:

wherein

-   R³ is a hydrogen atom or a halogen atom,-   m is 0 or 1, and-   * is a binding site to X or Y.-   [11] The method according to any of the above-mentioned [1] to [8],    wherein the aryl boronic acid compound represented by the    formula (II) is 4-boronophenylalanine,    4-borono-2-fluorophenylalanine or 3-boronophenylalanine, and the    radiolabeled aryl compound represented by the formula (I) is    4-astato(²¹¹At)phenylalanine, 4-astato(²¹¹At)-2-fluorophenylalanine    or 3-astato(²¹¹At)phenylalanine.-   [12] 3-Astato(²¹¹At)phenylalanine or a salt thereof.

Effect of the Invention

According to the production method of the present invention, theradiolabeled aryl compound (I) can be easily produced in a highradiochemical yield in a short time, and the formulation can be carriedout immediately after the labelling. Therefore, the labelling andformulation can be easily carried out in a short time, and a sequence ofprocesses from preparation of a radionuclide to RI internal therapy ordiagnosis for cancer can be promptly carried out.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows a thin layer chromatography (TLC) of the aqueous ²¹¹Atsolution prepared in Reference Example 1.

FIG. 2 shows a thin layer chromatography (TLC) of the reaction solutionof Example 1.

FIG. 3 shows a thin layer chromatography (TLC) of the reaction solutionof Example 2.

FIG. 4 shows a thin layer chromatography (TLC) of the reaction solutionof Example 3.

FIG. 5 shows a thin layer chromatography (TLC) of the reaction solutionof Example 4.

FIG. 6 a shows a thin layer chromatography (TLC) of the aqueous Na¹²³Isolution, and FIG. 6 b shows a thin layer chromatography (TLC) of thereaction solution of Example 5.

FIG. 7 shows a cellulose acetate membrane electrophoresis of thereaction solution of Example 6.

FIG. 8 shows a thin layer chromatography (TLC) of the reaction solutionof Example 7.

FIG. 9 shows comparison of 4-²¹¹At-Phe or 3-²¹¹At-Phe amount taken up byC6 glioma derived from rat in Example 10.

FIG. 10 shows SPECT imaging of C6 glioma-transplanted rat by 4-²¹¹At-Phein Example 11 (30 minutes and 3 hours after administration).

DESCRIPTION OF EMBODIMENTS

The present invention is explained in detail in the following.

In the present specification, examples of the “C₆₋₁₄ aryl group” includephenyl, 1-naphthyl, 2-naphthyl, 1-anthryl, 2-anthryl and 9-anthryl.

In the present specification, examples of the “halogen atom” include afluorine atom, a chlorine atom, a bromine atom and an iodine atom.

In the present specification, examples of the “C₁₋₆ alkyl group” includemethyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl,tert-butyl, pentyl, isopentyl, neo-pentyl, 1-ethylpropyl, hexyl,isohexyl, 1,1-dimethylbutyl, 2,2-dimethylbutyl, 3,3-dimethylbutyl and2-ethylbutyl.

Each symbol in the formulas (I) and (II) is explained below.

In the formulas (I) and (II), Ar is a C₆₋₁₄ aryl group optionally havingsubstituent(s).

The “C₆₋₁₄ aryl group” of the “C₆₋₁₄ aryl group optionally havingsubstituent(s)” represented by Ar is preferably phenyl.

Examples of the “substituent” of the “C₆₋₁₄ aryl group optionally havingsubstituent(s)” represented by Ar include groups capable of bindingspecifically to a target molecule. Examples of the target moleculeinclude antigens, transporters, receptors, enzymes, genes and the like,which are specifically expressed or overexpressed in cancer cells.Specific examples of such “substituent” include C₁₋₆ alkyl groups(preferably methyl, ethyl) substituted by a carboxy group and an aminogroup; a carboxy group; an amino group; a guanidino group; groups havinga tropane skeleton; fatty-acid residues (groups obtained by removing anyone hydrogen atom from fatty-acids); residues of biologically relatedsubstances such as peptides, proteins, antibodies, nucleic acids and thelike (groups obtained by removing any one hydrogen atom frombiologically related substances); and the like.

Ar is preferably a C₆₋₁₄ aryl group having substituent(s), morepreferably a phenyl group having substituent(s), still more preferably aresidue derived from an amino acid having phenyl group(s), or a residuederived from a peptide having phenyl group(s).

As used herein, the above-mentioned “residue derived from an amino acidhaving phenyl group(s)” means a group obtained by removing, from anamino acid having phenyl group(s) (e.g., phenylalanine or phenylglycineoptionally substituted by halogen atom(s), etc.), any one hydrogen atomon the phenyl ring.

Preferable example is a group represented by the formula:

wherein

-   R² is a halogen atom,-   m is 0 or 1,-   n is 0 or an integer of 1 to 4, and-   * is a binding site to X or Y.

More preferable example is a group represented by the formula:

wherein

-   R³ is a hydrogen atom or a halogen atom,-   m is 0 or 1, and-   * is a binding site to X or Y.

In another embodiment, more preferable example is a group represented bythe formula:

wherein

-   R³ is a hydrogen atom or a halogen atom,-   m is 0 or 1, and-   * is a binding site to X or Y.

The above-mentioned “residue derived from a peptide having phenylgroup(s)” means a group obtained by removing, from a peptide havingphenyl group(s) (e.g., a peptide containing phenylalanine orphenylglycine optionally substituted by halogen atom(s), etc.), any onehydrogen atom on the phenyl ring.

Preferable example is a group derived from a peptide having a partialstructure represented by the formula:

wherein

-   R² is a halogen atom,-   m is 0 or 1,-   n is 0 or an integer of 1 to 4, and-   * is a binding site to X or Y.

More preferable example is a group derived from a peptide having apartial structure represented by the formula:

wherein

-   R³ is a hydrogen atom or a halogen atom,-   m is 0 or 1, and-   * is a binding site to X or Y.

In another embodiment, more preferable example is a group derived from apeptide having a partial structure represented by the formula:

wherein

-   R³ is a hydrogen atom or a halogen atom,-   m is 0 or 1, and-   * is a binding site to X or Y.

The “halogen atom” represented by R² or R³ is preferably a fluorineatom.

R² is preferably a fluorine atom.

n is preferably 0 or 1.

-   R³ is preferably a hydrogen atom or a fluorine atom.-   m is preferably 1.

In the formula (II), Y is a borono group (—B(OH)₂) or an ester groupthereof.

Examples of the “ester group of borono group” represented by Y includethe following ester groups.

wherein R¹ is a C₁₋₆ alkyl group,

Y is preferably a borono group (—B(OH)₂).

In the formula (I) , X is a radionuclide ²¹¹At, ²¹⁰At, ¹²³I, ¹²⁴I, ¹²⁵Ior ¹³¹I.

When the radiolabeled aryl compound (I) or aryl boronic acid compound(II) is in the form of a salt, examples of such salts include metalsalts (e.g., alkali metal salts such as sodium salt, potassium saltetc.; alkaline-earth metal salts such as calcium salt, magnesium salt,barium salt etc.), an ammonium salt, salts with an organic base (e.g.,trimethylamine, triethylamine, pyridine, picoline, 2,6-lutidine), saltswith an inorganic acid (e.g., hydrochloric acid, hydrobromic acid,nitric acid, sulfuric acid), salts with an organic acid (e.g., formicacid, acetic acid, trifluoroacetic acid, phthalic acid, fumaric acid,oxalic acid, tartaric acid, maleic acid, citric acid, succinic acid,malic acid), and the like.

In the present invention, the radiolabeled aryl compound (I) is producedby reacting the aryl boronic acid compound (II) with a radionuclideselected from ²¹¹At, ²¹⁰At, ¹²³I, ¹²⁴I, ¹²⁵I and ¹³¹I in the presence ofan oxidizing agent selected from an alkali metal iodide, an alkali metalbromide, N-bromosuccinimide, N-chlorosuccinimide and hydrogen peroxide,in water.

The aryl boronic acid compound (II) is preferably an amino acid having aborono-substituted phenyl group(s), or a peptide containing the aminoacid, more preferably an amino acid having a borono-substituted phenylgroup(s). The borono-substituted phenyl group optionally has additionalsubstituent(s) such as a halogen and the like.

The aryl boronic acid compound (II) is more preferably4-boronophenylalanine, 4-borono-2-fluorophenylalanine,4-boronophenylglycine or 4-borono-2-fluorophenylglycine, particularlypreferably 4-boronophenylalanine or 4-borono-2-fluorophenylalanine.

In another embodiment, the aryl boronic acid compound (II) is morepreferably 4-boronophenylalanine, 4-borono-2-fluorophenylalanine,4-boronophenylglycine, 4-borono-2-fluorophenylglycine,3-boronophenylalanine or 3-boronophenylglycine, particularly preferably4-boronophenylalanine, 4-borono-2-fluorophenylalanine or3-boronophenylalanine.

The aryl boronic acid compound (II) is not limited to the compoundsexemplified above, and the radiolabelling method of aryl compound of thepresent invention can also be applied to various aryl boronic acidcompounds, for example, boronohydroxybenzene and boronocarboxybenzene(carboxyphenylboronic acid).

When the aryl boronic acid compound (II) is the above-mentioned aminoacid having a borono-substituted phenyl group(s), it is used generallyin the form of an aqueous solution, preferably in the form of a solutiondissolved in an aqueous alkali solution such as an aqueous sodiumhydrogencarbonate solution and the like.

Examples of the alkali metal iodide include sodium iodide, potassiumiodide and the like. Among them, preferred is sodium iodide.

Examples of the alkali metal bromide include sodium bromide, potassiumbromide and the like. Among them, preferred is sodium bromide.

The combination of the radionuclide and the oxidizing agent ispreferably

-   (1) a combination of the radionuclide of ²¹¹At or ²¹⁰At, and the    oxidizing agent selected from sodium iodide, sodium bromide,    N-bromosuccinimide, N-chlorosuccinimide and hydrogen peroxide; or-   (2) a combination of the radionuclide of ¹²³I, ¹²⁴I, ¹²⁵I or ¹³¹I,    and the oxidizing agent selected from N-bromosuccinimide and    N-chlorosuccinimide. The oxidizing agent may be used alone or in    combination of two or more kinds thereof. The oxidizing agent is    used generally in the form of an aqueous solution.

The oxidizing agent is used in an amount sufficient to oxidize theradionuclide, generally in a large excess amount relative to theradionuclide. It is used preferably in a concentration of 0.0001 to 0.2mol/L, more preferably in a concentration of 0.001 to 0.1 mol/L, interms of reaction efficiency and economic efficiency.

The radionuclide is used in the reaction generally in the form of anaqueous solution, preferably in the form of a solution prepared bydissolving in an aqueous alkali solution such as an aqueous sodiumhydrogencarbonate solution and the like, in terms of stability.

In cases where the radionuclide is ²¹¹At, first, bismuth is irradiatedwith helium particles accelerated to 28 MeV by cyclotron, and ²¹¹At isgenerated by the resulting ²⁰⁹Bi(α,2n)²¹¹At nuclear reaction. Next, byheating, the target substance ²⁰⁹Bi is melted, but the ²¹¹At isvaporized, and then the vaporized ²¹¹At is trapped in liquid nitrogen,and dissolved in water to prepare an ²¹¹At undiluted solution. Then, forthe purpose of stabilization of ²¹¹At, an aqueous alkali solution suchas an aqueous sodium hydrogencarbonate solution and the like is added tothe undiluted solution to prepare an aqueous ²¹¹At alkali solution.

In cases where the radionuclide is ²¹⁰At, first, bismuth is irradiatedwith helium particles accelerated to 29 MeV or more by cyclotron, and²¹⁰At is generated by the resulting ²⁰⁹Bi(α,3n)²¹⁰At nuclear reaction.Next, by the same procedure mentioned above, an aqueous ²¹⁰At solutionis prepared.

In cases where the radionuclide is ¹²³I, it is available as an aqueousNa¹²³I solution.

In cases where the radionuclide is ¹²⁴I first, tellurium is irradiatedwith proton particles accelerated by cyclotron, and ¹²⁴I is generated bythe resulting ¹²⁴Te(p,n)¹²⁴I nuclear reaction. Next, the targetsubstance ¹²⁴Te is melted, and the remaining ¹²⁴I is dissolved in anaqueous sodium hydroxide solution to prepare an aqueous ¹²⁴I sodiumhydroxide solution.

In cases where the radionuclide is ¹²⁵I, it is available as an aqueousNa¹²⁵I solution.

In cases where the radionuclide is ¹³¹I, it is available as an aqueousNa¹³¹I solution.

Since ²¹¹At has a short half-life of 7.2 hours, ²¹⁰At has a shorthalf-life of 8.3 hours, and ¹²³I has a short half-life of 13.2 hours,these radionuclides should be used in the subsequent reactionimmediately after the preparation. While ¹²⁴I has a relatively longhalf-life of 4.2 days, ¹²⁵I has a relatively long half-life of 59.4days, and ¹³¹I has a relatively long half-life of 8.04 days, theseradionuclides are also preferably used in the subsequent reactionimmediately after the preparation.

The aryl boronic acid compound (II) is used generally in a large excessamount relative to the radionuclide, preferably in a concentration of0.0001 mol/l to 0.5 mol/l, more preferably in a concentration of 0.001mol/l to 0.2 mol/l, per 1 Bq to 1,000 GBq of the radionuclide, in termsof reaction efficiency and economic efficiency.

The above-mentioned reaction is carried out by mixing the aryl boronicacid compound (II), an oxidizing agent and a radionuclide, and themixing order is not particularly limited. The reaction is preferablycarried out by adding an aqueous radionuclide alkali solution and anaqueous oxidizing agent solution, in this order, to an aqueous solution(preferably an aqueous sodium hydrogencarbonate solution) of the arylboronic acid compound (II), or by adding an aqueous oxidizing agentsolution and an aqueous radionuclide alkali solution, in this order, toan aqueous solution (preferably an aqueous sodium hydrogencarbonatesolution) of the aryl boronic acid compound (II), more preferably byadding an aqueous radionuclide alkali solution and an aqueous oxidizingagent solution, in this order, to an aqueous solution (preferably anaqueous sodium hydrogencarbonate solution) of the aryl boronic acidcompound (II).

The above-mentioned reaction is carried out in water, i.e., in anorganic solvent-free system.

The above-mentioned reaction is carried out at room temperature,specifically at 0° C.-40° C., preferably 10° C.-35° C. In the productionmethod of the present invention, the reaction proceeds rapidly in ashort time, even at room temperature. For example, the reaction iscompleted for 1 minute to 3 hour, particularly 1 minute to 30 minutes.

The completion of the reaction is confirmed by thin layer chromatography(TLC) analysis, based on the disappearance of a free radionuclide,

In the production method of the present invention, the radiolabeled arylcompound (I) can be obtained in a high radiochemical yield of 75% ormore, particularly 80% or more, especially 90% or more.

Since the reaction solution contains neither an organic solvent nor atoxic reagent, the reaction solution can be formulated into an injectionand the like immediately after the completion of the reaction, withoutisolation of the radiolabeled aryl compound (I).

As explained above, in the production method of the present invention,the labeling can be easily carried out in a high radiochemical yield ina short time, without use of an organic solvent and a toxic reagent.Therefore, a sequence of processes from preparation of a radionuclide toRI internal therapy or diagnosis for cancer can be promptly carried out.

The radiolabeled aryl compound (I) produced by such method is preferably

-   4-astato(²¹¹At)phenylalanine,-   3-astato(²¹¹At)phenylalanine,-   4-astato(²¹¹At)-2-fluorophenylalanine,-   4-astato(²¹⁰At)phenylalanine,-   3-astato(²¹⁰At)phenylalanine,-   4-astato(²¹⁰At)-2-fluorophenylalanine,-   4-iodo(¹²³I)phenylalanine,-   3-iodo(¹²³I)phenylalanine,-   4-iodo(¹²³I)-2-fluorophenylalanine,-   4-iodo(¹²⁴I)phenylalanine,-   3-iodo(¹²⁴I)phenylalanine,-   4-iodo(¹²⁴I)-2-fluorophenylalanine,-   4-iodo(¹²⁵I)phenylalanine,-   3-iodo(¹²⁵I)phenylalanine,-   4-iodo(¹²⁵I)-2-fluorophenylalanine,-   4-iodo(¹³¹I)phenylalanine,-   3-iodo(¹³¹I)phenylalanine,-   4-iodo(¹³¹I)-2-fluorophenylalanine,-   4-astato(²¹¹At)phenylglycine,-   3-astato(²¹¹At)phenylglycine,-   4-astato(²¹¹At)-2-fluorophenylglycine,-   4-astato(²¹⁰At)phenylglycine,-   3-astato(²¹⁰At)phenylglycine,-   4-astato(²¹⁰At)-2-fluorophenylglycine,-   4-iodo(¹²³I)phenylglycine,-   3-iodo(¹²³I)phenylglycine,-   4-iodo(¹²³I)-2-fluorophenylglycine,-   4-iodo(¹²⁴I)phenylglycine,-   3-iodo(¹²⁴I)phenylglycine,-   4-iodo(¹²⁴I)-2-fluorophenylglycine,-   4-iodo(¹²⁵I)phenylglycine,-   3-iodo(¹²⁵I)phenylglycine,-   4-iodo(¹²⁵I)-2-fluorophenylglycine,-   4-iodo(¹³¹I) phenylglycine,-   3-iodo(¹³¹I)phenylglycine, or-   4-iodo(¹³¹I)-2-fluorophenylglycine,-   more preferably-   4-astato(²¹¹At)phenylalanine,-   3-astato(²¹¹At)phenylalanine,-   4-astato(²¹¹At)-2-fluorophenylalanine,-   4-astato(²¹⁰At)phenylalanine,-   3-astato(²¹⁰At)phenylalanine,-   4-astato(²¹⁰At)-2-fluorophenylalanine,-   4-iodo(¹²³I)phenylalanine,-   3-iodo(¹²³I)phenylalanine,-   4-iodo(¹²³I)-2-fluorophenylalanine,-   4-iodo(¹²⁴I)phenylalanine,-   3-iodo(¹²⁴I)phenylalanine,-   4-iodo(¹²⁴I)-2-fluorophenylalanine,-   4-iodo(¹²⁵I)phenylalanine,-   3-iodo(¹²⁵I)phenylalanine,-   4-iodo(¹²⁵I)-2-fluorophenylalanine,-   4-iodo(¹³¹I)phenylalanine,-   3-iodo(¹³¹I)phenylalanine, or-   4-iodo(¹³¹I)-2-fluorophenylalanine,-   particularly preferably-   4-astato(²¹¹At)phenylalanine,-   3-astato(²¹¹At)phenylalanine, or-   4-astato(211At)-2-fluorophenylalanine.

Among the radiolabeled aryl compound (I), 3-astato(²¹¹At)phenylalanineis a novel compound. The compound is taken up in large amounts by cancercells, and therefore, it is particularly expected to be applied to RIinternal therapy for cancer.

EXAMPLES

The present invention is explained in detail by referring to thefollowing Examples, which are not to be construed as limitative, and theinvention may be changed within the scope of the present invention.

In the following Examples and Reference Examples, the radiochemicalyield is calculated by the following formula. radiochemical yield(%)=(radioactivity of the desired compound on thin-layer plate orelectrophoretic membrane/total radioactivity on thin-layer plate orelectrophoretic membrane)×100

The thin-layer plate and electrophoretic membrane was exposed on BASimaging plate (GE Healthcare), and the BAS imaging plate was analyzed byan image analyzer (Tyhoon FLA7000, GE Healthcare). The data processingwas performed using ImageQuantTL Analysis Toolbox (GE Healthcare).

Reference Example 1 Preparation of an Aqueous ²¹¹At Solution

²¹¹At was generated by ²⁰⁹Bi(α,2n)²¹¹At nuclear reaction, irradiatingbismuth with helium particles accelerated (28 MeV) by cyclotron. Afterthe irradiation, by heating, the target substance ²⁰⁹Bi was melted, butthe ²¹¹At was vaporized, and then the vaporized ²¹¹At was trapped inliquid nitrogen, and dissolved in a small amount of water to give an²¹¹At undiluted solution. To the obtained ²¹¹At undiluted solution wasadded a 7% aqueous sodium hydrogencarbonate solution to prepare anaqueous ²¹¹At solution having a radioactive concentration of about 5MBq/ml (immediately after the production). The thin layer chromatography(TLC) of the aqueous ²¹¹At solution are shown in FIG. 1 (thin-layerplate: G60 (Merck), developing solvent: ACN:water:TFA (66:33:1)). Thespots of ²¹¹At were detected on Rf=1.0 (80%) and 0.89 (8%).

Example 1 Synthesis of 4-²¹¹At-L-phenylalanine (Oxidizing Agent: NCS)

4-Borono-L-phenylalanine (Bpa) was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared a 10 mg/ml of solution. Thesolution (0.2 ml) was put into a small glass vial, and the aqueous ²¹¹Atsolution (5 MBq/ml, 0.2 ml) prepared in Reference Example 1 was addedthereto, and then an aqueous N-chlorosuccinimide (NCS) solution (4mg/ml, 0.04 ml) was slowly added dropwise thereto at room temperature.After 30 minutes, the reaction solution was analyzed by thin layerchromatography (TLC) (thin-layer plate: G60 (Merck), developing solvent:ACN:water:TFA (66:33:1)) (FIG. 2 ). The radiochemical yield of4-²¹¹At-L-phenylalanine (Rf=0.73) was 95%.

Example 2 Synthesis of 4-²¹¹At-L-phenylalanine (Oxidizing Agent: NBS)

4-Borono-L-phenylalanine (Bpa) was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared a 10 mg/ml of solution. Thesolution (0.2 ml) was put into a small glass vial, and the aqueous ²¹¹Atsolution (5 MBq/ml, 0.2 ml) prepared in Reference Example 1 was addedthereto, and then an aqueous N-bromosuccinimide (NBS) solution (4 mg/ml,0.04 ml) was slowly added dropwise thereto at room temperature. After 30minutes, the reaction solution was analyzed by thin layer chromatography(TLC) (thin-layer plate: G60 (Merck), developing solvent: ACN:water:TFA(66:33:1)) (FIG. 3 ). The radiochemical yield of 4-²¹¹At-L-phenylalanine(Rf=0.68) was 75.3%.

Example 3 Synthesis of 4-²¹¹At-L-phenylalanine (Oxidizing Agent: NaI)

4-Borono-L-phenylalanine (Bpa) was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared a 10 mg/ml of solution. Thesolution (0.2 ml) was put into a small glass vial, and the aqueous ²¹¹Atsolution (5 MBq/ml, 0.2 ml) prepared in Reference Example 1 was addedthereto, and then an aqueous sodium iodide (NaI) solution (10 mg/ml, 0.1ml) was slowly added dropwise thereto at room temperature. After 30minutes, the reaction solution was analyzed by thin layer chromatography(TLC) (thin-layer plate: G60 (Merck), developing solvent: ACN:water:TFA(66:33:1)) (FIG. 4 ). The radiochemical yield of 4-²¹¹At-L-phenylalanine(Rf=0.79) was 90%.

Example 4 Synthesis of 4-²¹¹At-L-phenylalanine (Oxidizing Agent: NaBr)

4-Borono-L-phenylalanine (Bpa) was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared a 10 mg/ml of solution. Thesolution (0.2 ml) was put into a small glass vial, and the aqueous ²¹¹Atsolution (5 MBq/ml, 0.2 ml) prepared in Reference Example 1 was addedthereto, and then an aqueous sodium bromide (NaBr) solution (10 mg/ml,0.1 ml) was slowly added dropwise thereto at room temperature. After 30minutes, the reaction solution was analyzed by thin layer chromatography(TLC) (thin-layer plate: G60 (Merck), developing solvent: ACN:water:TFA(66:33:1)) (FIG. 5 ). The radiochemical yield of 4-²¹¹At-L-phenylalanine(Rf=0.63) was 84.7%.

Example 5 Synthesis of 4-¹²³I-L-phenylalanine (Oxidizing Agent: NBS)

4-Borono-L-phenylalanine (Bpa) was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared a 10 mg/ml of solution. Thesolution (0.2 ml) was put into a small glass vial, and an aqueous Na¹²³Isolution (74 MBq/ml, 5 mmol/l aqueous NaOH solution, 0.2 ml) was addedthereto, and then an aqueous N-bromosuccinimide (NBS) solution (4 mg/ml,0.04 ml) was slowly added dropwise thereto at room temperature. After 30minutes, the reaction solution was analyzed by thin layer chromatography(TLC) (thin-layer plate: G60 (Merck), developing solvent: ACN:water:TFA(66:33:1)) (FIG. 6 b ). The radiochemical yield of4-¹²³I-L-phenylalanine (Rf=0.70) was 93%. The thin layer chromatography(TLC) (thin-layer plate: G60 (Merck), developing solvent: ACN:water:TFA(66:33:1)) of the aqueous Na¹²³I solution is shown in FIG. 6 a.

Reference Example 2 Synthesis of 4-iodo-L-phenylalanine UsingNon-Radioactive Iodine

4-Borono-L-phenylalanine (Bpa) was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared a 10 mg/ml of solution. Thesolution (0.3 ml) was put into a small glass vial, and an aqueous NaIsolution (10 mg/ml, 0.3 ml) was added thereto, and then an aqueousN-bromosuccinimide (NBS) solution (10 mg/ml, 0.3 ml) was slowly addeddropwise thereto at room temperature. After 30 minutes, the reactionsolution was analyzed by thin layer chromatography (TLC) (thin-layerplate: G60 (Merck), developing solvent: ACN:water:TFA (66:33:1)), andthe single spot was detected on Rf=0.7 (ultraviolet radiation andcoloration in iodine bath). Then, the reaction solution was 1000fold-diluted with water, and analyzed by LC-MS for amino acid analysis,compared to commercially available 4-iodo-L-phenylalanine as a controlsample. The product was detected at the same retention time as in thecontrol sample 4-iodo-L-phenylalanine, and the extraction mass was alsothe same as that of the control sample (theoretical mass=290.9756,extraction mass=291.9835). The purity of the product was 98.9%. Theimpurity was L-phenylalanine (1.1%) alone, and 4-bromo-L-phenylalanineand the like were not detected.

Example 6 Synthesis of 4-²¹¹At-2-fluoro-L-phenylalanine (OxidizingAgent: NBS)

4-Borono-2-fluoro-L-phenylalanine (FBpa) was dissolved in a 7% aqueoussodium hydrogencarbonate solution to prepared a 5 mg/ml of solution. Thesolution (0.2 ml) was put into a small glass vial, and the aqueous ²¹¹Atsolution (5 MBq/ml, 0.1 ml) prepared in Reference Example 1 was addedthereto, and then an aqueous N-bromosuccinimide (NBS) solution (4 mg/ml,0.04 ml) was slowly added dropwise thereto at room temperature. After 15minutes, the reaction solution was analyzed by cellulose acetatemembrane electrophoresis method (FIG. 7 ). The radiochemical yield of4-²¹¹At-2-fluoro-L-phenylalanine (Rf=0.68) was 92.2%.

Example 7 Synthesis of 3-²¹¹-At-D,L-phenylalanine

3-Borono-D,L-phenylalanine (3-Bpa) was dissolved in a 1.4% aqueoussodium hydrogencarbonate solution to prepared a 10 mg/ml of solution.The solution (0.2 ml) was put into a small glass vial, and the aqueous²¹¹At solution (5 MBq/ml, 0.2 ml) prepared in Reference Example 1 wasadded thereto, and then an aqueous N-bromosuccinimide (NBS) solution (4mg/ml, 0.04 ml) was slowly added dropwise thereto at room temperature.After 30 minutes, the reaction solution was analyzed by thin layerchromatography (TLC) (thin-layer plate: silica gel G60 (Merck),developing solvent: ACN:water:TFA (66:33:1)) (FIG. 8 ). Theradiochemical yield of the produced 3-²¹¹At-D,L-phenylalanine (Rf=0.76)was 93.3%.

Example 8 1-²¹¹At-2-hydroxybenzene

2-Boronohydroxybenzene was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared an aqueous solution having aconcentration of 5 mg/ml. The aqueous solution was put into a smallglass vial, and the aqueous ²¹¹At solution (5 MBq/ml, 0.2 ml) preparedin Reference Example 1 was added thereto, and then an aqueousN-bromosuccinimide (NBS) solution (4 mg/ml, 0.04 ml) was slowly addeddropwise thereto at room temperature. After 30 minutes, an aqueousascorbic acid solution (3 mg/ml, 0.03 ml) was added to the reactionsolution to quench the reaction. The reaction solution was analyzed bythin layer chromatography (TLC) (thin-layer plate: silica gel G60(Merck), developing solvent: ACN:water:TFA (66:33:1)). The radiochemicalyield of the produced 1-²¹¹At-2-hydroxybenzene (Rf=0.91) was 98.1%. Thisresult demonstrates that the method of radiolabelling an aryl compoundof the present invention can be applied not only to boronophenylalaninebut also to boronohydroxybenzene.

Example 9 1-²¹¹At-4-carboxybenzene

4-Barboxyphenylboronic acid was dissolved in a 7% aqueous sodiumhydrogencarbonate solution to prepared an aqueous solution having aconcentration of 16 mg/ml. The aqueous solution was put into a smallglass vial, and the aqueous ²¹¹At solution (5 MBq/ml, 0.2 ml) preparedin Reference Example 1 was added thereto, and then an aqueousN-bromosuccinimide (NBS) solution (4 mg/ml, 0.04 ml) was slowly addeddropwise thereto at room temperature. After 30 minutes, an aqueousascorbic acid solution (3 mg/ml, 0.03 ml) was added to the reactionsolution to quench the reaction. The reaction solution was analyzed bythin layer chromatography (TLC) (thin-layer plate: silica gel G60(Merck), developing solvent: ACN:waterTFA (66:33:1)). The radiochemicalyield of the produced 1-²¹¹At-4-carboxybenzene (Rf=0.81) was 87.5%. Thisresult demonstrates that the method of radiolabelling an aryl compoundof the present invention can be applied not only to boronophenylalaninebut also to boronocarboxybenzene (carboxyphenylboronic acid).

Example 10 Glioma Cell Uptake Experiment

The concentration of the 4-²¹¹At-L-phenylalanine (4-²¹¹At-Phe) solutionprepared in Example 2 and the 3-²¹¹At-D,L-phenylalanine (3-²¹¹At-Phe)solution prepared in Example 7 were each adjusted to 5 MBq/2 mg·ml. C6glioma cell derived from rat was seeded on each well of 24-well plate by5×10⁵ cells/well, and the cells were used in the next day's experiment(cell number: about 1×10⁶ cells/well). Before the experiment, the mediumwas replaced with an amino acid-free HESS medium (0.5 ml/well).4-²¹¹At-Phe or 3-²¹¹At-Phe was added to each well of the plate by 10 μl,and the wells were divided into three groups of inhibitor-free group, 1%Phe addition group (competitive inhibitor) and 100 mM BCH addition group(LAT1 inhibitor), and incubated at 37° C. for 30 minutes or 60 minutes.After the incubation, the culture solution was removed, and the cellswere washed with PBS, and lysed, and the amount of radioactivity takenup intracellularly was measured. The results are shown in FIG. 9 .4-²¹¹At-Phe and 3-²¹¹At-Phe were both taken up by the cell, and the celluptake of 3-²¹¹At-Phe was slightly more than that of 4-²¹¹At-Phe.Moreover, the cell uptake after 60 minutes was reduced, compared to thatafter 30 minutes. The competitive inhibitor (Phe) reduced the celluptake of 4-²¹¹At-Phe and 3-²¹¹At-Phe to ½ to ⅓, and LAT1 inhibitor(BCH) reduced the cell uptake of 4-²¹¹At-Phe and 3-²¹¹At-Phe to ¼ to ⅛.That is to say, the results demonstrated that 4-²¹¹At-Phe and3-²¹¹At-Phe were both specifically taken up by the cells via LAT1, andtherefore, 4-²¹¹At-Phe and 3-²¹¹At-Phe can be expected to be applied toRI internal therapy for cancer. Especially, the cell uptake of3-²¹¹At-Phe was more than that of 4-²¹¹At-Phe, and therefore,3-²¹¹At-Phe can be further expected for the above-mentioned application.

Example 11 SPECT Imaging of C6 Glioma-Transplanted Rat by4-²¹¹At-L-phenylalanine

The 4-²¹¹At-L-phenylalanine (4-At-Phe) (1 MBq) prepared in Example 2 wasadministered to the tail vein of the glioma-transplanted rat, and therat was imaged by SPECT camera (E-cam, Siemens). The SPECT images 30minutes and 3 hours after the administration are shown in FIG. 10 . Inboth images, the accumulation of 4-²¹¹At-Phe in the tumor (both flanks,indicated by arrows in the figure) was observed. Therefore, 4-²¹¹At-Phecan be expected to be applied to RI internal therapy or diagnosis forcancer.

INDUSTRIAL APPLICABILITY

According to the production method of the present invention, theradiolabeled aryl compound (I) can be easily produced in a highradiochemical yield in a short time, and the formulation can be carriedout immediately after the labelling. Therefore, the labelling andformulation can be easily carried out in a short time, and a sequence ofprocesses from preparation of a radionuclide to RI internal therapy ordiagnosis for cancer can be promptly carried out.

This application is based on patent application No. 2017-151632 filed onAug. 4, 2017 in Japan, the contents of which are encompassed in fullherein.

The invention claimed is:
 1. A method of producing a radiolabeled arylcompound represented by formula (I):Ar—X   (I) wherein Ar is a C₆₋₁₄ aryl group optionally havingsubstituent(s), and X is ²¹¹At or ²¹⁰At, or a salt thereof, whichcomprises reacting an aryl boronic acid compound represented by formula(II):Ar—Y   (II) wherein Ar is as defined above, and Y is a borono group(—B(OH)₂) or an ester group thereof, or a salt thereof, with aradionuclide selected from ²¹¹At and ²¹⁰At, in the presence of an agentselected from the group consisting of an alkali metal iodide and analkali metal bromide in water, in an organic solvent-free system.
 2. Themethod according to claim 1, wherein the reaction is carried out at roomtemperature.
 3. The method according to claim 1, wherein Y is a boronogroup (—B(OH)₂).
 4. The method according to claim 1, wherein thesubstituent of the C₆₋₁₄ aryl group optionally having substituent(s)represented by Ar is a group capable of binding specifically to a targetmolecule.
 5. The method according to claim 4, wherein the targetmolecule is an antigen, a transporter, a receptor, an enzyme or a gene,which target molecule is specifically expressed or overexpressed in acancer cell.
 6. The method according to claim 1, wherein Ar is a grouprepresented by formula:

wherein R² is a halogen atom, m is 0 or 1, n is 0 or an integer of 1 to4, and * is a binding site to X or Y, or a group derived from a peptidehaving a partial structure represented by formula:

wherein R² is a halogen atom, m is 0 or 1, n is 0 or an integer of 1 to4, and * is a binding site to X or Y.
 7. The method according to claim1, wherein Ar is a group represented by formula:

wherein R³ is a hydrogen atom or a halogen atom, m is 0 or 1, and * is abinding site to X or Y, or a group derived from a peptide having apartial structure represented by formula:

wherein R³ is a hydrogen atom or a halogen atom, m is 0 or 1, and * is abinding site to X or Y.
 8. The method according to claim 1, wherein thearyl boronic acid compound represented by formula (II) is4-boronophenylalanine, 4-borono-2-fluorophenylalanine or3-boronophenylalanine, and the radiolabeled aryl compound represented byformula (I) is 4-astato(²¹¹ At)phenylalanine, 4-astato(²¹¹At)-2-fluorophenylalanine or 3-astato(²¹¹ At)phenylalanine. iodide. 9.The method according to claim 1, wherein the agent is an alkali metaliodide.
 10. The method according to claim 1, wherein the agent is analkali metal bromide.
 11. The method according to claim 9, X is ²¹¹At.12. The method according to claim 9, X is ²¹⁰At.
 13. The methodaccording to claim 10, X is ²¹¹At.
 14. The method according to claim 10,X is ²¹⁰At.